Precise synaptic connectivity is essential for the proper function of the nervous system. Surprisingly, however, we know very little about mechanisms that regulate the formation of neuron-neuron synapses in vivo. This application proposes to use the simple and accessible peripheral synapse in the mouse superior cervical ganglion (SCG) as a model system to study the formation of neuron-neuron synapses. The focus of this proposed research is to determine key molecular mechanisms that regulate the assembly of the postsynaptic apparatus at neuronal cholinergic synapses. Neuronal cholinergic systems play critical roles in both the peripheral and central nervous systems. Alterations in cholinergic function have been implicated in several neurological disorders, including Alzheimer' s disease and schizophrenia. Currently, mechanisms regulating the formation, maturation and stability of neuronal cholinergic synapses are completely unknown. Here, we propose to use molecular and genetic approaches to elucidate the mechanisms that regulate the assembly of the postsynaptic complex at neuronal cholinergic synapses. The first aim is to identify the molecular components of the postsynaptic signaling complex at neuronal cholinergic synapses and to understand how the postsynaptic signaling complex is assembled. The second aim is to determine the molecular mechanisms that target/anchor neuronal nicotinic acetylcholine receptors to synapses. Finally, we will use mutant mice to test the in vivo function of one postsynaptic signaling complex protein we already identified at the SCG synapse and assess the importance of the postsynaptic signaling complex in the formation, maturation and stability of neuronal cholinergic synapses. These studies will advance our understanding of how the precise synaptic connectivity in the nervous system is established and regulated.